Saliva as a biomarker for head and neck squamous cell carcinoma: In vitro detection of cytotoxic effects by using the plating efficiency index

Abstract

Abstract. Worldwide, the incidence of head and neck squamous cell carcinoma (HNSCC) is increasing. The development of an effective prevention program would provide a promising opportunity to control this disease. We applied the 'plating efficiency index' of Chinese hamster lung fibroblasts to demonstrate the presence of cytotoxic effects in saliva samples from cancer patients as well as from healthy probands. Correlations between individual risk factors and the cytotoxic effects of saliva specimens were analysed and evaluated. Saliva samples were obtained from male patients (n=43) with carcinomas of the upper aerodigestive tract, and from a healthy age-matched male control group (n=131) with different tobacco and alcohol consumption habits. The extraction of non-stimulated sober saliva was chosen for this investigation. In vitro cytotoxicity of the saliva was detected using the permanent mammalian cell line V79 (lung fibroblasts) of Chinese hamsters. The determination of the plating efficiency index was made in the logarithmic growth phase of the initial cell culture. A reduction in cell colonies (plating efficiency) of at least 50% was defined as strongly cytotoxic. A significant direct correlation was found between daily tobacco consumption and the reduction of plating efficiency (p<0.0001). We found only a moderate increase in the cytotoxicity of tested saliva samples in correlation with daily alcohol uptake. The difference between tumor patients and healthy probands was highly significant (χ 2 test; p=0.001). Using the method of logistic regression analysis, we found a 3.6-fold increased cancer risk in probands with cytotoxic saliva (p<0.001). According to our results, the plating efficiency index seems to be a suitable method for the detection of increased cancer risk. In combination with several effective biomarkers on cytogenetic end-points, it may help to establish biomonitoring programs for secondary cancer prevention. IntroductionPrimary cancers of the head and neck region have varying types of histology. Squamous cell carcinomas (SCCs) are among the most common malignancies of the upper aerodigestive tract (1). The incidence of these tumors varies worldwide on the basis of changing 'lifestyle' factors (2,3) such as chronic tobacco and alcohol consumption or changing exposure to environmental carcinogens. Despite multimodal therapies with often mutilating surgical procedures (especially in extended tumor stages), the prognosis for these tumors remains poor. Hence, the development of effective prevention programs would provide a promising opportunity to control this increasingly common disease.In general, there are two different methods of disease prevention. First, and most effective, is the reduction of avoidable exogenous risk factors. The second involves the identification of people bearing individual increased cancer risk, which may help in the detection of early-stage carcinomas (4-7). Hopefully, this will be achieved in the future through the use of biomarkers showing biological effects, like per...

Salivary VEGF: a non-invasive angiogenic and lymphangiogenic proxy in head and neck cancer prognostication

“…The relevance of this pilot study lies in its possible development as a prognostic test with utility in the development of an effective prevention program to control head and neck cancer [ 8 ]. When compared with other highly diagnostic salivary tumour biomarkers, the advantages of this simple ELISA over the laboratory based 'in vitro detection of cytotoxic effects of saliva by using the plating efficiency index' [ 9 ] lie mainly in the ease of collection, simplicity of use and speed of results of the former. Despite this both tests have significant merit and may in fact be complementary.…”

Are oral and dental diseases linked to cancer?

“…The authors concluded that polymicrobial supragingival plaque may be a co‐factor in the development of oral carcinomas. The same authors, using a cell culture model, observed that saliva from patients with head and neck tumors had a higher cytotoxic effect on fibroblasts in comparison with saliva from healthy controls (Bloching et al , 2007a,b). Hence, the mutagenicity of saliva needs to be taken into account when considering the mechanisms involved in oral carcinomas.…”

“…Saliva may also contain mutagenic compounds. Bloching et al (2007a,b) reported that a high dental plaque index and large number of decayed teeth were significantly correlated with Salmonella typhimurium genotoxicity (Ames test). The authors concluded that polymicrobial supragingival plaque may be a co‐factor in the development of oral carcinomas.…”

Applicability of non-invasively collected matrices for human biomonitoring

“…Additionally, saliva specimens have been used in combination with the Ames test or a Chinese hamster V79 lung fibroblast cell line to investigate the genotoxic effects of smoking and alcohol consumption. exfoliated buccal cells have been used to monitor genetic damage in humans using the micronucleus test [ 81 , 82 ].…”